The objective of this proposal is to develop an innovative technique to increase both the detection efficiency and mass resolution of large DNA ions when matrix-assisted laser desorption/ionization (MALDI) is used for DNA sequencing. MALDI based time-of-flight mass spectrometry (TOFMS) has been proven to be capable of fast sequencing of small DNAs using an ion detector. The time needed for the sequencing is less than 1 ms. However, the sensitivity of ion detection critically depends on the speed of the ions. High-speed ions strike the detector's surface and eject secondary electrons. The electrons are multiplied and the ion signal is amplified. The detection efficiency drops rapidly with the increase of molecular mass since the speed of large DNA is lower than that of the small DNA. Due to the low detection efficiency for the large DNA, a high laser pumping intensity for ablation is used. The high pumping intensity creates complicated collision and energy transfer processes in MALDI and results in a poor mass resolution in sequencing large DNA. The investigators propose to improve the ion detection efficiency by using UV laser-induced photodissociation/fragmentation at the end of the drift tune of a TOFMS. The large parent DNA ions interact with and are fragmented into small ions by a UV laser. The small fragments of DNA ions are then mass-selected and accelerated by pulsed high voltage (HV) electric fields to a speed higher than the critical speed needed for efficient ejection of secondary electron from the surface of an ion detector. With special care being taken, the detection sensitivity of large DNA sequencing is expected to be improved by several orders of magnitude. The improved sensitivity would make possible the application of MALDI-based TOFMS for routine sequencing of very large DNA (m/z-1,000,000-Dalton or higher) and allow the use of lower laser intensity for ablation, which consequently results in a better mass resolution.